Two highly-immunogenic recombinant Env polypeptides of the HIV-1 gp41 (protein 566) and HIV-2 gp35 (protein 996) transmembrane glycoproteins were expressed in E. coli in quantity, purified in milligram quantities to near homogeneity, and used in immunoassays (Western and dot blots) against sera from Central and West Africa, Mexico, and the United States to distinguish between HIV-1 and HIV-2 infections. In addition, recombinant HTLV-I (rpBI) and HTLV-II (rpBII) gp46 Env polypeptides, and HTLV-I p2lE Env polypeptides (protein 400), previously expressed in E. coli, were also used in immunoassays (Western blot) to distinguish between HTLV-I and HTLV-ll infections. Moreover, antibodies developed to the HTLV-I (rpBI) and HTLVII (rpBII) Env polypeptides were used in antibody-dependent cellular cytotoxicity (ADCC) assays to delineate the type-specific epitopes residing on the surfaces of the HTLV-I and HTLV-II Env glycoproteins. Currently available serological tests do not distinguish between HIV-1 and HIV-2 or HTLV-I and HTLV-ll infections because of significant antigen-antibody crossreactivity due to the close genetic relatedness between HIV-l/HIV-2 and HTLVI/HTLV-II virus groups. Thus, the application of these highly-immunogenic HIV-1 (protein 566), HIV-2 (protein 996), HTLV-I (rpBI and protein 400) and HTLV-II (rpBII) candidate antigens in development of immunoassays for differentiating single, as well as dual viral infections, and as potential subunit vaccines, requires further serological and biological characterization.